Liquid chromatography (LC) columns have been extensively developed and are used routinely in both analytical and preparative chromatography. The separation in a liquid chromatography column of a sample comprising a mixture of components is achieved by conveying the sample in a liquid mobile phase through a stationary phase, thereby causing the sample to separate into its components due to different partitioning between the mobile and stationary phases of each of the components (i.e. the components have different partition coefficients). In liquid chromatography the stationary phase is typically in the form of a bed of particles packed within the column, which is usually a tubular column. This invention relates to such so-called packed columns wherein the column media comprises packed particles.
Silica particles are commonly used as the stationary phase bed although other materials may be used. Non-porous particles have a low sample capacity. Therefore, porous particles are commonly used which contain a network of pores to increase the surface area of the stationary phase and thus improve the capacity of the separation. Larger porous particles tend to have longer mass transfer distances for the sample, which leads to slower separation and broader retention time peaks than with smaller porous particles. Consequently, there has been a trend to reduce the size of the porous particles in order to improve the kinetics and resolution of the separation. This is important for separation of large biomolecules, such as proteins for example. However, a consequence of using smaller porous particles is an increased resistance to flow and therefore a higher operating pressure is needed in order to deliver the required flow. Higher pressures increase the cost and complexity of the instrumentation.
An alternative to the aforementioned fully porous particles, in which the pores extend throughout the bulk of the particles, has been the use of so-called fused core particles, which are also sometimes termed superficially porous particles. These are particles that have a non-porous core (also termed a fused or solid core) and are only porous in an outer layer or region (also termed a shell) that surrounds the core, i.e. they are only porous at the surface, not throughout the bulk of the particles. These particles have some of the advantages of small porous particles, such as short mass transfer distances provided by the outer porous shell and hence high chromatographic efficiency, but the overall particle size is larger due to the solid core to enable lower operating pressures to be used. Fused core particles also offer the possibility of having narrower particle size distributions than porous particles due to the ability to classify larger, heavier particles more effectively.
Although the use of fused core particles has improved the column efficiency that can be delivered with lower pressures, there nevertheless remains a need to reduce still further the operating pressure whilst maintaining good column efficiency.
Against this background the present invention has been made.